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| United States Patent |
6,258,572
|
|
Giselbrecht
, et al.
|
July 10, 2001
|
Process for preparing L-aspartic acid
Abstract
An improved process for preparing L-aspartic acid by enzyme-catalyzed
reaction of fumaric acid with ammonia, in which process L-aspartic acid is
precipitated out by nitric acid and the resultant mother liquor is
subjected to a nanofiltration.
| Inventors:
|
Giselbrecht; Karl-Heinz (Pasching, AT);
Schaller; Josef (Linz, AT)
|
| Assignee:
|
DSM Fine Chemicals Austria Nfg GmbH & CoKG (Linz, AT)
|
| Appl. No.:
|
315044 |
| Filed:
|
May 20, 1999 |
Foreign Application Priority Data
| Current U.S. Class: |
435/109; 435/232 |
| Intern'l Class: |
C12P 013/20 |
| Field of Search: |
435/109,232
|
References Cited
U.S. Patent Documents
| 5952206 | Sep., 1999 | Giselbrecht et al. | 435/109.
|
| 6150142 | Nov., 2000 | Mukouyama et al. | 435/109.
|
| Foreign Patent Documents |
| 298438 | May., 1972 | AT.
| |
| 0 798 377 | Oct., 1997 | EP.
| |
Other References
Chemical Abstracts, 238:74403s (1998).
|
Primary Examiner: Lilling; Herbert J.
Attorney, Agent or Firm: Wenderoth, Lind & Ponack, L.L.P.
Claims
What is claimed is:
1. A process for preparing L-aspartic acid by enzyme-catalyzed reaction of
fumaric acid with ammonia, which comprises
a) reacting fumaric acid with ammonia to form ammonium L-aspartate in the
presence of aspartase or aspartase-producing microorganisms in an inert
diluent,
b) removing any excess ammonia from the reaction mixture and
c) precipitating L-aspartic acid by adding nitric acid, separating it off
from the reaction mixture, washing and drying it, then
d) separating off residual L-aspartic acid and residual fumaric acid from
the precipitation filtrate by nanofiltration, recycling the retentate to
step c) and
e) concentrating the filtrate from step d), adding the resultant distillate
in step a) to any ammonia excess separated off in step b) and feeding the
concentrated filtrate to a nitrogen fertilizer process.
2. The process as claimed in claim 1, wherein the diluent used in step a)
is water, water/ethanol or water/acetone mixtures.
3. The process as claimed in claim 1, wherein step a) is carried out at a
pH between 7 and 9.
4. The process as claimed in claim 1, wherein, in step b), a distillation
is carried out at temperatures between 30 and 110.degree. C. and at
atmospheric pressure or under reduced pressure to remove said excess
ammonia.
5. The process as claimed in claim 1, wherein step c) is carried out at a
pH between 2 and 5.
6. The process as claimed in claim 1, wherein step c) is carried out at a
temperature between 15 and 60.degree. C.
7. The process as claimed in claim 1, wherein the nanofiltration in step d)
is carried out at a pH between 4 and 11, at a pressure between 1 and 50
bar and at a temperature between 10 and 50.degree. C.
8. The process as claimed in claim 7, wherein the pH is between 7 and 10.
9. The process as claimed in claim 1, wherein the filtrate from step d) is
concentrated to an above 50 to 80% strength solution.
Description
L-Aspartic acid is an essential starting material for a wide variety of
additives for the pharmaceutical industry and the food sector. For
example, L-aspartic acid is used for preparing artificial sweetener, such
as aspartame. Therefore, a multiplicity of chemical and enzymatic
processes have already been described for preparing L-aspartic acid. In
the enzymatic variants, L-aspartic acid is usually obtained by enzymatic
addition of ammonia to fumaric acid with subsequent precipitation from the
resultant ammonium L-aspartate solution.
According to the prior art, L-aspartic acid can be precipitated, for
example, by adding sulfuric acid or hydrochloric acid or other acids, such
as p-toluenesulfonic acid.
However, the disadvantages of this are that the loss of ammonia is high and
a large amount of wastewater having a high concentration of ammonium salts
of the acids used is discharged.
For this reason, attempts have been made to find possible methods of
decreasing or completely avoiding the wastewater problem.
According to U.S. Pat. No. 4,560,653, the L-aspartic acid is precipitated,
for example, by adding maleic acid. After separating off L-aspartic acid,
the remaining mother liquor is subjected to an isomerization step in which
maleic acid is isomerized into fumaric acid, for instance using a catalyst
comprising bromine ions, then purified and fed back to the enzymatic
reaction. In order to bypass the isomerization step, other suitable
additives for precipitating L-aspartic acid have been sought.
In the Japanese Laid-Open Application JP 08-33493 (Chem. Abstracts 1224:315
167), the use of fumaric acid or salt of fumaric acid as precipitant is
described. The disadvantage of this process variant is the poor water
solubility of fumaric acid, as a result of which, during the workup of the
mother liquor, either large amounts of water have to be distilled off or a
very dilute procedure having high reaction volumes is necessary.
The object of the invention is therefore to find a process which avoids the
previous problems in the precipitation of L-aspartic acid and leads to
L-aspartic acid in high yields and high purity.
Unexpectedly, this object was achieved by using nitric acid as precipitant
and subsequent nanofiltration of the mother liquor.
The invention therefore relates to an improved process for preparing
L-aspartic acid by enzyme-catalyzed reaction of fumaric acid with ammonia,
which comprises
a) reacting fumaric acid with ammonia to form ammonium L-aspartate in the
presence of aspartase or aspartase-producing microorganisms in an inert
diluent,
b) removing any excess ammonia from the reaction mixture and
c) precipitating L-aspartic acid by adding nitric acid, separating it off
from the reaction mixture, washing and drying it, then
d) separating off residual L-aspartic acid and residual fumaric acid from
the precipitation filtrate by nanofiltration, recycling the retentate to
step c) and
e) concentrating the filtrate from step d), adding the resultant distillate
in step a) to any ammonia excess separated off in step b) and feeding the
concentrated filtrate to a nitrogen fertilizer process.
In the process according to the invention, L-aspartic acid is precipitated
by nitric acid. The reaction streams are recirculated according to the
invention. The process according to the invention can be seen in FIG. 1,
where the reaction streams are shown diagrammatically.
The concentrated filtrate obtained in step e) can be introduced directly
into the nitrogen fertilizer process.
The first step of the process according to the invention comprises the
enzymatic reaction of fumaric acid with ammonia. The reaction takes place
here in an inert diluent. Suitable inert diluents are water, water/ethanol
or water/acetone mixtures and the like. Preferably, water is used. Fumaric
acid can be used here in a concentration of up to the solubility limit, so
that either a solution or a suspension is obtained. Into this solution or
suspension is introduced ammonia, gaseous, liquefied or in the form of a
from 10 to 35% strength by weight solution, as a result of which the
temperature increases to 60.degree. C. and a pH of between 7 and 9 is
established.
Preferably, an aqueous 20 to 30% strength by weight ammonia solution is
used. Into the resultant system, preferably a solution, is then stirred
the 20 enzyme aspartase, or an aspartase-producing micro-organism, at from
20 to 60.degree. C., preferably at from 30 to 50.degree. C. In this
addition of enzyme- or aspartase-producing microorganism, it is
advantageous if the addition of ammonia gives a solution, since in the
case of a suspension more enzyme is necessary, owing to adsorption of the
enzyme and loss in activity caused by this. For a virtually quantitative
reaction after up to from 24 to 30 hours, in this case, from 30 to 50 IU
(enzyme activity) are required per mole of fumaric acid.
Aspartase-producing microorganisms are, for example, Pseudomonas
fluorescens, Protens vulgaris, Pseudomonas aeruginosa, Serratia
marcescens, Bacterium succinium, Bacillus subtilis, Aerobacter aerogenes,
Micrococcus sp., Escherichia coli, inter alia.
Other suitable aspartase-producing microorganisms are described, for
example, in U.S. Pat. No. 3,791,926 and U.S. Pat. No. 3,198,712.
In the process according to the invention, in addition, purified or
synthetic aspartase can be used. The enzyme or the aspartase-producing
microorganism can be added in liquid or immobilized form, as described,
for example, in EP 0 127 940.
After completion of the reaction, the end of the reaction can be determined
photometrically, for example, according to step b), if appropriate, the
ammonia is separated off by distillation or stripping.
Step b) can be carried out at atmospheric pressure or under reduced
pressure and at temperatures of from 30 to 110.degree. C., preferably from
40 to 90.degree. C.
Excess ammonia is removed from the reaction mixture in this case using
familiar distillation methods, for example using short-path or thin-film
evaporators, strippers etc. According to the distillation method, either,
preferably, atmospheric pressure, or reduced pressure between 80 and 200
mbar is employed. The resultant ammonia distillate is reused as starting
material for a subsequent further enzyme reaction a) together with the
distillate from the subsequent process step d).
After the removal of ammonia, or directly after step a), the L-aspartic
acid is precipitated as in step c) For this purpose, the ammonium
L-aspartate solution is admixed with sufficient nitric acid to achieve a
pH between 2 and 5, preferably up to the isoelectric point. Nitric acid is
added in this case preferably as from 10 to 65% strength, particularly
preferably as 60% strength, nitric acid. The temperature of the reaction
mixture in this case is between 15 and 60.degree. C., preferably between
20 and 35.degree. C.
From the 2nd reaction cycle, in addition to the nitric acid, the retentate
from the nanofiltration of the precipitation filtrate produced in the
subsequent step d), which retentate comprises residual amounts of
L-aspartic acid and fumaric acid, is added to the ammonium L-aspartate
solution.
The reaction mixture is then cooled, preferably to 0 to -15.degree. C. and
the L-aspartic acid which has crystallized out is separated off, for
example by filtration, such as by absorption filtration or by
centrifugation. Preferably, L-aspartic acid is separated off by
centrifugation or decanting.
The L-aspartic acid crystals filtered off are finally washed, preferably
with water, and dried. The wash water can here be recycled to step a) or,
preferably, subjected to the subsequent nanofiltration. The remaining
precipitation filtrate produced after separating off L-aspartic acid, as
well as the wash water are then subjected to a nanofiltration in step d).
The pH in this case is preferably between 4 and 11, particularly
preferably between 7 and 10, and is preferably set using NH.sub.3. The
pressure is preferably between 1 and 50 bar. The temperature is preferably
10 to 50.degree. C., particularly preferably 15 to 30.degree. C. Owing to
the nanofiltration, the L-aspartic acid present in the filtrate, as well
as residual amounts of fumaric acid as ammonium salts which are present in
the retentate can be recovered from the filtrate. The resultant retentate
is fed to the crystallization process in step c) from the 2nd reaction
cycle.
The filtrate produced by the nanofiltration (approximately 20% strength
ammonium nitrate solution) is concentrated in step e), for example by
evaporation of water, until an over 50 to 80% strength, preferably an
approximately 60 to 70% strength, ammonium nitrate solution is obtained.
This solution can, as an ammonium nitrate solution free of organic carbon,
be introduced into the nitrogen fertilizer process directly as material of
value or starting material. The distillate remaining from the
concentration is, from the 2nd cycle, together with any ammonia excess
separated off in step b), added to the enzyme-catalyzed reaction in step
a).
By means of the process according to the invention, L-aspartic acid can be
obtained in quantitative yields, i.e. up to 99% and a content of >99.5%.
EXAMPLE 1
Into 850 ml (950 g) of L-aspartic acid reaction solution (DSM-Chemie Linz),
comprising 240 g (1.8 mol) of L-aspartic acid, prepared by reaction from
210 g (1.8 mol) of fumaric acid, 200 ml of 25% strength by weight ammonia
solution in 280 ml of H.sub.2 O, in the presence of 0.08 ml of aspartase
solution (1100 IU/ml) was added sufficient 65% strength nitric acid to
reach pH 2.7 (see Table 1). After 115 minutes, the mixture was cooled to
0.degree. C. by ice, the sediment which had precipitated out was
centrifuged off, rinsed with 100 ml of distilled H.sub.2 O and centrifuged
dry for 10 minutes. 312 g of solid having a moisture content of 22.9% were
obtained. This is equivalent to 225.2 g (i.e. 93.9%) of anhydrous
L-aspartic acid.
In addition, 123.7 g (124 ml) of wash water and 834.5 g (800 ml) of
precipitation filtrate (mother liquor M1) were obtained. The wash water
and M1 were adjusted to pH 9 using 25% strength ammonia water and then
subjected to a nanofiltration. The roughly 20% ammonium nitrate solution
thus obtained was concentrated to 65%. The distillate was added back to
the crystallization process in later batches in step c).
TABLE 1
Time min g HNO.sub.3 ml HNO.sub.3 pH Temp.
0 0 0 10.2 23.4
10 71 49 9.0 30.5
20 88 63 8.0 29.8
25 95 65 6.0 30.0
35 117 84 5.0 30.1
40 135 98 4.5 to 5.4 30.1 to 31.3
55 150 108 5.4 30.6
65 186 133 5.0 32.6
90 229 166 4.5 32.2
100 247 178 4.0 32.5
105 255 184 3.5 32.4
110 259 186 3.0 32.2
115 262 190 2.7 31.6
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